Neutron Reflection Study of Surface Adsorption of Fc, Fab, and the Whole mAb
Neutron Reflection Study of Surface Adsorption of Fc, Fab, and the Whole mAb
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DOI：
10.1021/acsami.7b06131
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发表时间：
2017-07-12
影响因子：
9.5
通讯作者：
Lu, Jian Ren
中科院分区：
文献类型：
--
作者：
Li, Zongyi;Li, Ruiheng;Lu, Jian Ren
表征片段结晶（Fc）和抗原结合片段（Fab）对单克隆抗体（mAb）在空气/水界面吸附的影响，是了解单克隆抗体液体制剂在生产、运输和储存过程中的稳定性的重要一步。在这里，中子反射被用来研究单抗及其Fc和Fab片段在空气/水中的吸附。通过改变同位素对比，可以明确地确定吸附层的吸附量、厚度、取向和浸没度。Fc吸附在1小时内达到饱和，其表面吸附量随体积浓度变化不大。相比之下，Fab的吸附速度较慢，吸附量与浓度有关。与Fab相比，高得多的Fc吸附与其较低的表面电荷有关。单抗吸附的时间和浓度依赖主要是Fab行为，尽管Fab和Fc行为都对单抗吸附量有贡献。将pH从5.5改变到8.8对Fc、Fab或mAb的吸附量没有太大影响。然而，在pH值为8-8.8时，Fc的吸附量略有下降，而Fab和mAb的吸附量反之亦然，这与Fab的优势行为一致。当体积浓度从S增加到50 ppm时，Fc层的厚度在40埃时几乎不变，而Fab和mAb层的厚度从45埃增加到50埃。这些结果表明，吸附的mAb、Fc和Fab均保留了它们的球状结构，并以垂直于界面的短轴向长度定向。
Characterizing the influence of fragment crystallization (Fc) and antigen-binding fragment (Fab) on monoclonal antibody (mAb) adsorption at the air/water interface is an important step to understanding liquid mAb drug product stability during manufacture, shipping, and storage. Here, neutron reflection is used to study the air/water adsorption of a mAb and its Fc and Fab fragments. By varying the isotopic contrast, the adsorbed amount, thickness, orientation, and immersion of the adsorbed layers could be determined unambiguously. While Fc adsorption reached saturation within the hour, its surface adsorbed amount showed little variation with bulk concentration. In contrast, Fab adsorption, was slower and the adsorbed amount was concentration dependent. The much higher Fc adsorption, as compared to Fab, was linked to its lower surface charge. Time and concentration dependence of mAb adsorption was dominated by Fab behavior, although both Fab and Fc behaviors contributed to the amount of mAb adsorbed. Changing the pH from 5.5 to 8.8 did not much perturb the adsorbed amount of Fc, Fab, or mAb. However, a small decrease in adsorption was obseived for the Fc over pH 8-8.8 and vice versa for the Fab and mAb, consistent with a dominant Fab behavior. As bulk concentration increased from S to 50 ppm, the thicknesses of the Fc layers were almost constant at 40 angstrom, while Fab and mAb layers increased from 45 to 50 angstrom. These results imply that the adsorbed mAb, Fc, and Fab all retained their globular structures and were oriented with their short axial lengths perpendicular to the interface.